Protein materials which are cross-linked to have a high molecular weight or gelled by the cross-linking method of the present invention can be used in the field of food processing such as of raw fish meat paste, kamaboko (fish cake), fish/livestock meat sausage, tofu (soy bean curd), noodles, confectionery/bread, food adhesives, sheet-like meat food, yogurt, jelly and cheese. In addition, they can also be used as novel protein-derived materials in a wide range of industries including cosmetics, raw materials of microcapsules and carriers of immobilized enzymes.
As enzymes having a possibility of increasing molecular weight of protein by cross-linking reaction, transglutaminase, lysyl oxidase, protein disulfide-isomerase, protein-disulfide reductase, sulfhydryl oxidase, lipoxygenase, polyphenol oxidase (tyrosinase) and peroxidase have been known (Matheis and Whitaker, J. Food Biochemistry, 11, 309-327, 1987).
Among the above-described enzymes having a possibility of increasing molecular weight of protein by cross-linking reaction, a method for cross-linking protein by transglutaminase is well known. It is also known that this method has been broadly used mainly in the field of food processing based on the discovery of an inexpensive microbial transglutaminase which does not require the presence of calcium for the reaction (JP-B-6-65280 (the term "JP-B" as used herein means an "examined Japanese patent publication"), Agric. Biol. Chem., vol. 69, no. 10, pp. 1301-1308).
The protein cross-linking reaction by transglutaminase, however, has the following problem. That is, since transglutaminase is an enzyme which forms an intramolecular or intermolecular bridge structure of protein as a result of the acyl rearrangement reaction generated between the .gamma.-carboxyl group of glutamine residue and the .epsilon.-amino group of lysine residue in a protein, some species of protein can hardly become the substrate for the enzyme due to insufficient glutamine residues or lysine residues. For example, albumin proteins cannot be used as the substrate for transglutaminase under their native state.
Thus, although a possibility of using several enzymes as the enzyme-aided protein cross-linking method has been indicated, most of them are not sufficiently useful in their supplying amounts, costs, easiness in the purification, and the like. Even if the microbial transglutaminase is used for the cross-linking method, its application is limited because of a problem that the reaction does not occur in some protein species. In consequence, great concern has been directed toward the development of a protein cross-linking method which uses other enzymes.